1. Field of the Invention
This invention relates generally to methods and compositions for the prevention and treatment of sepsis and particularly to the use of a combination of complement inhibitors and CD14 pathway inhibitors to prevent or treat sepsis.
2. Description of the Prior Art Complement
The immune system protects the body against pathogenic bacteria, viruses, parasites and other harmful organisms. The immune system is divided into two components, the humoral system and the cellular system. Generally, the humoral system includes the complement system and the production of antibodies to defend against pathogens. The complement system, or simply complement, involves the production of proteins that assist the antibodies in the host defense. Complement is a group of at least 30 surface-bound and soluble proteins. The activity of the soluble proteins is destroyed by heating serum at 56° C. for 30 minutes. Complement proteins are involved in the opsonization of microorganisms for phagocytosis, direct killing of microorganisms by lysis, chemotactic attraction of leukocytes to sites of inflammation, activation of leukocytes, and processing of immune complexes.
Complement proteins work in a cascade wherein the binding of one protein promotes the binding of the next protein in the cascade. Activation of the cascade leads to release of biologically active small peptides called anaphylatoxins (C3a, C4a, and the most potent C5a) contributing to the inflammatory reaction, and eventually in the formation of a membrane attack complex (C5b-9 or MAC) that may lyse the target cell. Different complement molecules are synthesized by different cell types, e.g. fibroblasts and intestinal epithelial cells make C1, while most of the components are synthesized in the liver.
The components and mechanism of the complement system are well known. Basically, there are three complement pathways, the classical pathway, the lectin pathway, and the alternative pathway. The classical pathway is triggered primarily by immune complexes containing antigen and IgG or IgM, but also by other agents like C-reactive protein. The lectin pathway is triggered by binding of mannose binding lectin (MBL) or ficolins to carbohydrate structures (e.g. mannan) on foreign surfaces. The alternative pathway is activated principally by repeating polysaccharides and other polymeric structures such as those found on bacteria.
The classical pathway is activated when the globular domains of C1q (part of the C1qrs complex) bind to the Fc fragments of IgM or multiple molecules of IgG. In the presence of calcium ions, this binding causes the autocatalytic activation of two C1r molecules. The C1r molecules activate two molecules of C1s. C1s is a serine protease that cleaves C4a from C4b. C4b immediately binds to adjacent proteins or carbohydrates on the surface of the target cell and then binds to C2 in the presence of magnesium ions. C1s cleaves C2b from this complex, yielding the classical pathway C3 convertase, C4b2a The C3 convertase cleaves many hundreds of molecules of C3 into C3a and C3b. Some molecules of C3b will bind back to C4b2a to yield the classical pathway C5 convertase, C4b2a3b. C5 convertase cleaves C5 into C5a and C5b. C5b binds to the surface of the cell, initiating the formation of MAC.
C3a, C4a, and C5a are all anaphylatoxins. C3a and C5a are also chemoattractants. C3a and C5a have the ability to bind to mast cells and basophils. C5a is also a potent activator of neutrophils, basophils and macrophages and causes induction of adhesion molecules on vascular endothelial cells. C5a also down regulates neutrophils and monocytes. When C3a and C5a bind their receptors on the mast cells and basophils, these cells release histamine and other highly active peptides into blood and tissues. These peptides increase the permeability of the vascular walls allowing neutrophils to migrate into the area Neutrophils are further encouraged to migrate to the site of complement activation due to the potent chemotactic (attractant) effect of C5a. The neutrophils phagocytose invading pathogens and also release mediators that attract macrophages to the site of infection. These cells also have the ability to phagocytose invading cells and further promote the inflammatory response and effectively eliminate many of the infections microorganisms.
The “lectin pathway” is similar to the classical pathway except it is initiated by the calcium-dependent lectin MBL that binds to terminal mannose groups on the surface of bacteria. MBL is analogous to C1q. When MBL binds to its target, it releases and thus activates three associated serine proteases known as MASP1, MASP2 and MASP3 (mannose-binding lectin-associated serine protease), which are analogous to C1r and C1s. Among them, MASP2 plays the key role in cleaving C4 into C4b and C4a and C2 into C2b and C2a Following the activation of C4 and C2, the lectin pathway is identical to the classical pathway.
The alternative complement pathway involves an amplification loop utilizing C3b produced by the classical pathway. Some molecules of C3b generated by the classical pathway C3 convertase are funneled into the alternative pathway. Surface-bound C3b binds Factor B to yield C3bB, which becomes a substrate for Factor D. Factor D is a serine protease that cleaves the Ba fragment, leaving C3bBb bound to the surface of the target cell. C3bBb is stabilized by properdin (P), forming the complex C3bBbP, which acts as the alternative pathway C3 convertase. As in the classical pathway, the C3 convertase participates in an amplification loop to cleave many C3 molecules, resulting in the deposition of C3b molecules on the target cell. Some of these C3b molecules bind back to C3bBb to form C3bBb3b, the alternative pathway C5 convertase. C5 convertase cleaves C5 into C5a and C5b. C5b binds to the surface of the cell to initiate the formation of the membrane attack complex.
The classical, lectin, and alternative pathways all end with the formation of C5 convertase. C5 convertase leads to the assembly of the MAC via the lytic pathway. Components C5-C8 attach to one another in tandem and promote the insertion of one or more monomers of C9 into the lipid bilayer of the target cell. This insertion leads to the formation of pores that cause calcium influx with subsequent cellular activation of nucleated cells or cell lysis and death if the attack is sufficiently strong.